vdsstream VDS - Target - 1-deoxy-D-xylulose-5-phosphate reductoisomerase (P. falciparum)

Target (protein/gene name): 1-deoxy-D-xylulose 5-phosphate
NCBI Gene # or RefSeq#: 812223 __https://www.ncbi.nlm.nih.gov/gene/812223__
Protein ID (NP or XP #) or Wolbachia#: __**XP_001348815.1**__
Organism: Plasmodium falciparum 3D7
Etiologic Risk Group (see link below):
Background/Disease Information:
Essentiality of this protein:
"PF14_0641 has essentiality data
Gene/Ortholog: mtu2917 (OG4_13658); Phenotype: non-essential; Source study: nmpdr
Gene/Ortholog: eco174 (OG4_13658); Phenotype: undefined; Source study: blattner
Gene/Ortholog: eco174 (OG4_13658); Phenotype: essential; Source study: gerdes
Gene/Ortholog: eco174 (OG4_13658); Phenotype: essential; Source study: keio
Gene/Ortholog: eco174 (OG4_13658); Phenotype: essential; Source study: shigen"
http://tdrtargets.org/targets/view?gene_id=2971

“Most significantly, the DOXP pathway has become attractive as a target for a new class of antimalarial agents since malaria parasites are efficiently killed in vitro and in the murine model by fosmidomycin and FR900098, both specific inhibitors of the DOXP reductoisomerase (Kuzuyama et al. 1998; Zeidler et al. 1998; Jomaa et al. 1999).”
Source: Plasmodium falciparum: Detection of the Deoxyxylulose 5-Phosphate Reductoisomerase Activity

Complex of enzymes: N/A
EC#: 1.1.1.26 Source: __http://tdrtargets.org/targets/view?gene_id=2971__
Link to BRENDA EC# page: http://www.brenda-enzymes.org/php/result_flat.php4?ecno=1.1.1.267

http://enzyme.expasy.org/EC/1.1.1.267

Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/General_Information/glyoxylate_reductase.Par.0001.File.tmp/glyoxylate_reductase.pdf
http://eprints.ru.ac.za/2712/1/TANNER-MSC-TR04-143.pdf
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC21430/?tool=pubmed

List cost and quantity of reagents and supplier (new item)

Reagent	Cost	Quantity	Product No.
Tris-HCl	$47.80	100g	T5941
MnCl2	$42.90	10g	244589
NADPH	$97.90	25mg	N5130

Supplied by Sigma-Aldrich

Structure Available (PDB or Homology model)
http://www.pdb.org/pdb/explore/explore.do?structureId=3AU8
http://www.pdb.org/pdb/explore/explore.do?structureId=3AU9
http://www.pdb.org/pdb/explore/explore.do?structureId=3AUA
Druggable Target (see Databases for this):
0.6
Source: http://tdrtargets.org/targets/view?gene_id=2971

Current Inhibitors:
“To date, several DXR inhibitors have been identified [19-23], fosmi- domycin and its analogue FR900098 have been shown to be the most potent...exhibit reduced pharmacological properties and their potential to become drugs is considered poor [24].”
Source: __http://benthamscience.com/ppl/sample/ppl17-1/0015E.pdf__

“...However, malarial recrudescence is a common problem because it is difficult to completely eradicate malaria parasites from the human body. Therefore, the refinement of the anti-malarial performance of fosmidomycin alone or in combination with another compound is expected to lead to more effective anti-malarial drugs.”
Source: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3216497/

Expression Information (has it been expressed in bacterial cells):
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2833050/?tool=pubmed

Purification Method:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2833050/?tool=pubmed

Image of protein (PyMol or etc):

Source: http://www.pdb.org/pdb/explore/explore.do?structureId=3AU8
Amino Acid Sequence:
1 mkkyiyiyff fititindlv inntskcvsi errknnayin ygigyngpdn kitksrrckr
61 iklckkdlid igaikkpinv aifgstgsig tnalniirec nkienvfnvk alyvnksvne
121 lyeqareflp eylcihdksv yeelkelvkn ikdykpiilc gdegmkeics snsidkivig
181 idsfqglyst myaimnnkiv alankesivs agfflkklln ihknakiipv dsehsaifqc
241 ldnnkvlktk clqdnfskin ninkiflcss ggpfqnltmd elknvtsena lkhpkwkmgk
301 kitidsatmm nkglevieth flfdvdyndi evivhkecii hscvefidks visqmyypdm
361 qipilysltw pdriktnlkp ldlaqvstlt fhkpslehfp ciklayqagi kgnfyptvln
421 asneiannlf lnnkikyfdi ssiisqvles fnsqkvsens edlmkqilqi hswakdkatd
481 iynkhnss
Source: https://www.ncbi.nlm.nih.gov/protein/XP_001348815.1

length of your protein in Amino Acids:
488 aa
Source: https://www.ncbi.nlm.nih.gov/protein/XP_001348815.1

Molecular Weight of your protein in kiloDaltons using the __**Expasy ProtParam**__ website
55756.7
Source: Expasy ProtParam
55757 DA
Source: http://tdrtargets.org/targets/view?gene_id=2971

Molar Extinction coefficient of your protein at 280 nm wavelength:
Ext. coefficient 48540 Abs 0.1% (=1 g/l) 0.871, assuming all pairs of Cys residues form cystines
Ext. coefficient 47790 Abs 0.1% (=1 g/l) 0.857, assuming all Cys residues are reduced

Source: Expasy ProtParam

CDS Gene Sequence:
ORIGIN
1 atgaagaaat atatttatat atattttttc ttcatcacaa taactattaa tgatttagta
61 ataaataata catcaaaatg tgtttccatt gaaagaagaa aaaataacgc atatataaat
121 tatggtatag gatataatgg accagataat aaaataacaa agagtagaag atgtaaaaga
181 ataaagttat gcaaaaagga tttaatagat attggtgcaa taaagaaacc aattaatgta
241 gcaatttttg gaagtactgg tagtataggt acgaatgctt taaatataat aagggagtgt
301 aataaaattg aaaatgtttt taatgttaaa gcattgtatg tgaataagag tgtgaatgaa
361 ttatatgaac aagctagaga atttttacca gaatatttgt gtatacatga taaaagtgta
421 tatgaagaat taaaagaatt ggtaaaaaat ataaaagatt ataaacctat aatattgtgt
481 ggtgatgaag ggatgaaaga aatatgtagt agtaatagta tagataaaat agttattggt
541 attgattctt ttcaaggatt atattctact atgtatgcaa ttatgaataa taaaatagtt
601 gcgttagcta ataaagaatc cattgtctct gctggtttct ttttaaagaa attattaaat
661 attcataaaa atgcaaagat aatacctgtt gattcagaac atagtgctat atttcaatgt
721 ttagataata ataaggtatt aaaaacaaaa tgtttacaag acaatttttc taaaattaac
781 aatataaata aaatattttt atgttcatct ggaggtccat ttcaaaattt aactatggac
841 gaattaaaaa atgtaacatc agaaaatgct ttaaagcatc ctaaatggaa aatgggtaag
901 aaaataacta tagattctgc aactatgatg aataaaggtt tagaggttat agaaacccat
961 tttttatttg atgtagatta taatgatata gaagttatag tacataaaga atgcattata
1021 cattcttgtg ttgaatttat agacaaatca gtaataagtc aaatgtatta tccagatatg
1081 caaataccca tattatattc tttaacatgg cctgatagaa taaaaacaaa tttaaaacct
1141 ttagatttgg ctcaggtttc aactcttaca tttcataaac cttctttaga acatttcccg
1201 tgtattaaat tagcttatca agcaggtata aaaggaaact tttatccaac tgtactaaat
1261 gcgtcaaatg aaatagctaa caacttattt ttgaataata aaattaaata ttttgatatt
1321 tcctctataa tatcgcaagt tcttgaatct ttcaattctc aaaaggtttc ggaaaatagt
1381 gaagatttaa tgaagcaaat tctacaaata cattcctggg ccaaagataa agctaccgat
1441 atatacaaca aacataattc ttcatag
//

Source: https://www.ncbi.nlm.nih.gov/nuccore/XM_001348779.1

GC% Content for gene: 24.88%
Source: http://bioinformatics.org/sms2/dna_stats.html
CDS Gene Sequence (codon optimized) - after having done Primer Design:
1 ATGAAAAAGTATATCTACATCTACTTCTTCTTTATCACTATCACGATTAACGACCTGGTC
61 ATTAACAACACGTCTAAATGCGTTTCTATTGAACGTCGTAAGAACAACGCTTACATCAAC
121 TACGGTATCGGTTACAACGGTCCAGATAATAAGATCACCAAGTCTCGTCGTTGCAAACGT
181 ATTAAACTGTGCAAAAAAGACCTCATCGATATTGGCGCGATCAAAAAGCCTATCAACGTA
241 GCGATCTTCGGTTCTACGGGCTCTATCGGTACCAACGCGCTCAACATCATTCGTGAATGT
301 AACAAAATTGAAAACGTCTTCAACGTGAAAGCGCTGTATGTAAATAAGTCTGTTAACGAG
361 CTGTACGAACAAGCCCGTGAGTTCCTGCCGGAATACCTGTGCATCCACGACAAAAGCGTT
421 TACGAAGAGCTGAAAGAACTGGTTAAAAACATCAAGGATTACAAGCCGATTATCCTGTGC
481 GGTGATGAAGGCATGAAGGAAATCTGCTCTTCTAATAGCATTGACAAGATCGTCATCGGT
541 ATTGACAGCTTCCAGGGTCTGTACTCTACCATGTACGCAATTATGAACAATAAGATTGTG
601 GCGCTGGCAAACAAAGAATCTATCGTTTCTGCGGGTTTTTTCCTCAAAAAACTGCTCAAT
661 ATCCACAAGAATGCCAAAATCATCCCGGTCGACTCTGAGCACTCTGCAATCTTCCAATGC
721 CTGGACAACAACAAGGTTCTCAAAACCAAGTGCCTCCAGGACAACTTCTCTAAAATCAAC
781 AATATCAATAAAATCTTTCTCTGCTCCTCTGGCGGTCCGTTCCAGAACCTGACCATGGAT
841 GAACTCAAGAACGTTACCTCTGAAAATGCCCTCAAGCACCCGAAGTGGAAAATGGGTAAG
901 AAGATTACTATCGATTCTGCGACCATGATGAATAAAGGTCTGGAAGTTATCGAAACCCAC
961 TTCCTGTTCGACGTTGACTACAATGACATTGAAGTAATCGTTCACAAAGAGTGCATTATC
1021 CACTCTTGCGTTGAATTTATCGACAAATCTGTTATTTCTCAGATGTACTACCCGGACATG
1081 CAGATCCCGATCCTGTATTCTCTGACCTGGCCGGACCGTATCAAGACCAATCTGAAACCG
1141 CTCGACCTCGCGCAGGTTTCTACGCTGACCTTCCACAAACCGTCTCTCGAACACTTCCCG
1201 TGCATCAAGCTGGCGTACCAGGCGGGTATTAAGGGTAACTTCTACCCGACCGTACTCAAC
1261 GCGTCCAACGAAATCGCGAATAACCTGTTTCTGAATAACAAGATTAAGTATTTCGACATC
1321 TCCTCTATCATCTCTCAGGTACTGGAGAGCTTCAACTCCCAGAAGGTTAGCGAGAATTCC
1381 GAAGACCTGATGAAACAAATCCTCCAGATTCACTCCTGGGCGAAAGACAAAGCGACTGAT
1441 ATCTATAACAAACACAACTCTTCTTGA

GC% Content for gene (codon optimized): 43.83%

Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
1 ATGAAAAAGTATATCTACATCTACTTCTTCTTTATCACTATCACGATTAACGACCTG 57
2 CGTTCAATAGAAACGCATTTAGACGTGTTGTTAATGACCAGGTCGTTAATCGTGATAGTG 60
3 GTCTAAATGCGTTTCTATTGAACGTCGTAAGAACAACGCTTACATCAACTACGGTATCGG 60
4 GACGAGACTTGGTGATCTTATTATCTGGACCGTTGTAACCGATACCGTAGTTGATGTAAG 60
5 GATAATAAGATCACCAAGTCTCGTCGTTGCAAACGTATTAAACTGTGCAAAAAAGACCTC 60
6 ACGTTGATAGGCTTTTTGATCGCGCCAATATCGATGAGGTCTTTTTTGCACAGTTTAATA 60
7 CGATCAAAAAGCCTATCAACGTAGCGATCTTCGGTTCTACGGGCTCTATCGGTACCAACG 60
8 GACGTTTTCAATTTTGTTACATTCACGAATGATGTTGAGCGCGTTGGTACCGATAGAGCC 60
9 GTGAATGTAACAAAATTGAAAACGTCTTCAACGTGAAAGCGCTGTATGTAAATAAGTCTG 60
10 GCAGGAACTCACGGGCTTGTTCGTACAGCTCGTTAACAGACTTATTTACATACAGCGCTT 60
11 AGCCCGTGAGTTCCTGCCGGAATACCTGTGCATCCACGACAAAAGCGTTTACGAAGAGCT 60
12 ATCGGCTTGTAATCCTTGATGTTTTTAACCAGTTCTTTCAGCTCTTCGTAAACGCTTTTG 60
13 ACATCAAGGATTACAAGCCGATTATCCTGTGCGGTGATGAAGGCATGAAGGAAATCTGCT 60
14 TGTCAATACCGATGACGATCTTGTCAATGCTATTAGAAGAGCAGATTTCCTTCATGCCTT 60
15 AGATCGTCATCGGTATTGACAGCTTCCAGGGTCTGTACTCTACCATGTACGCAATTATGA 60
16 ATAGATTCTTTGTTTGCCAGCGCCACAATCTTATTGTTCATAATTGCGTACATGGTAGAG 60
17 CGCTGGCAAACAAAGAATCTATCGTTTCTGCGGGTTTTTTCCTCAAAAAACTGCTCAATA 60
18 AGAGTCGACCGGGATGATTTTGGCATTCTTGTGGATATTGAGCAGTTTTTTGAGGAAAAA 60
19 AATCATCCCGGTCGACTCTGAGCACTCTGCAATCTTCCAATGCCTGGACAACAACAAGGT 60
20 TGATTTTAGAGAAGTTGTCCTGGAGGCACTTGGTTTTGAGAACCTTGTTGTTGTCCAGGC 60
21 TCCAGGACAACTTCTCTAAAATCAACAATATCAATAAAATCTTTCTCTGCTCCTCTGGCG 60
22 GTAACGTTCTTGAGTTCATCCATGGTCAGGTTCTGGAACGGACCGCCAGAGGAGCAGAGA 60
23 ATGGATGAACTCAAGAACGTTACCTCTGAAAATGCCCTCAAGCACCCGAAGTGGAAAATG 60
24 TTATTCATCATGGTCGCAGAATCGATAGTAATCTTCTTACCCATTTTCCACTTCGGGTGC 60
25 GATTCTGCGACCATGATGAATAAAGGTCTGGAAGTTATCGAAACCCACTTCCTGTTCGAC 60
26 ACTCTTTGTGAACGATTACTTCAATGTCATTGTAGTCAACGTCGAACAGGAAGTGGGTTT 60
27 ATTGAAGTAATCGTTCACAAAGAGTGCATTATCCACTCTTGCGTTGAATTTATCGACAAA 60
28 TCTGCATGTCCGGGTAGTACATCTGAGAAATAACAGATTTGTCGATAAATTCAACGCAAG 60
29 TACTACCCGGACATGCAGATCCCGATCCTGTATTCTCTGACCTGGCCGGACCGTATCAAG 60
30 CAGCGTAGAAACCTGCGCGAGGTCGAGCGGTTTCAGATTGGTCTTGATACGGTCCGGCCA 60
31 GCGCAGGTTTCTACGCTGACCTTCCACAAACCGTCTCTCGAACACTTCCCGTGCATCAAG 60
32 GTCGGGTAGAAGTTACCCTTAATACCCGCCTGGTACGCCAGCTTGATGCACGGGAAGTGT 60
33 ATTAAGGGTAACTTCTACCCGACCGTACTCAACGCGTCCAACGAAATCGCGAATAACCTG 60
34 AGAGGAGATGTCGAAATACTTAATCTTGTTATTCAGAAACAGGTTATTCGCGATTTCGTT 60
35 GATTAAGTATTTCGACATCTCCTCTATCATCTCTCAGGTACTGGAGAGCTTCAACTCCCA 60
36 GGATTTGTTTCATCAGGTCTTCGGAATTCTCGCTAACCTTCTGGGAGTTGAAGCTCTCCA 60
37 CGAAGACCTGATGAAACAAATCCTCCAGATTCACTCCTGGGCGAAAGACAAAGCGACTGA 60
38 TCAAGAAGAGTTGTGTTTGTTATAGATATCAGTCGCTTTGTCTTTCGC 48

Source:
LOGFILE.txt

Primer design results for 'tail' primers (this is just 2 sequences):

We designed a forward and reverse primer that will allow us to PCR the P.fal. gene and insert it into an expression vector (pNIC-Bsa4).

Forward Primer:
5’ TACTTCCAATCCATGAAAAAGTATATCTACATCTAC 3’ 36 bp
GC Content 30.6%
0 mM Mg2+ Tm 56.7 oC 1.5 mM Mg2+ Tm 64.9 oC 2 mM Mg2+ Tm 65.4 oC
4 mM Mg2+ Tm 66.5 oC 6 mM Mg2+ Tm 67.1 oC

Reverse Primer:
5’ AAACACAACTCTTCTTGA 3’ Reverse complement it:
5’ TATCCACCTTTACTGTCAAGAAGAGTTGTGTTT3’ 33 bp
GC Content 36.4%
0 mM Mg2+ Tm 59.7 oC 1.5 mM Mg2+ Tm 67.6 oC 2 mM Mg2+ Tm 68.1 oC
4 mM Mg2+ Tm 69.2 oC 6 mM Mg2+ Tm 69.7 oC

Transmembrane Protein Prediction (several algorithms):